The type 1 interferons (IFNs), including the IFNs alpha and beta, have potent antiviral effects, but also mediate a wide range of immunoregulatory functions. These include better-characterized effects on NK cells and incompletely understood effects on CD8 T cells. Some are paradoxical, and the mechanisms controlling type 1 IFN effects, to allow access to subsets when needed, are poorly characterized. The cytokines do stimulate a signaling pathway, depending on the signal transducers and activators of transcription (STAT) 1 and 2, to induce direct antiviral effects, but they can conditionally activate all of the STATs, including STAT4, and STAT4 contributes to IFN-? induction. As a result of experiments demonstrating that total STAT1 protein is dramatically induced at early times after infection, and that type 1 IFN activation of STAT4 negatively correlates with STAT1 levels, this project proposed to test the hypotheses that type 1 IFN effects are controlled by regulation of access to different intracellular signaling pathways, and that this regulation is required and delivered during innate and adaptive immune responses to viral infections. The work has proven the hypotheses to be correct. Unexpected mechanisms for the delivery of access to STAT1 as compared to STAT4 were discovered at the levels of differential expression of STAT4 and of apparent competition for binding to the type 1 IFN receptor, and these were shown to be important for health. Experiments are proposed in this renewal application to test the hypotheses that STATs act as master switches in promoting cellular responses by both enhancing and inhibiting type 1 IFN effects, and that the STAT1 and STAT4 effects on the reciprocal signaling pathway are influenced by, and have consequences for, other STAT molecules. This will be accomplished through four specific aims focusing on NK and CD8 T cell responses. Aim 1 will define the use of STATs as master switches to mediate positive and negative effects on responsiveness to type 1 IFNs. Aim 2 will mechanistically define the pathways regulating STAT levels. Aim 3 will broaden the characterization of type 1 IFN access to other STATs. Aim 4 will define the importance of modulating STAT levels and access for delivery of changing type 1 IFN biological functions during viral infection. Immunological, virological, biochemical, and molecular techniques will be used, and the work will be advanced by studies of responses ex vivo following exposure to cytokines, and in vivo following lymphocytic choriomeningitis virus infections in wild type and genetically altered mice. The project promises to continue to advance understanding of the control of type 1 IFN effects in shaping immune responses to infection, and to provide insights for use of cytokines in therapeutic applications. Even more broadly, however, it will identify paradigms for how cytokine effects are regulated to add value to a limited number of genes.